Browse
Submit Data
Databases
API
Help

Dataset Information

25 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Genetic manipulation of Staphylococcus aureus.

ABSTRACT: Staphylococcus aureus is a facultative anaerobic Gram-positive coccus and a member of the normal skin flora, as well as that of the nasal passages of humans. However, S. aureus can also gain entry into the host and cause life-threatening infections or persist as disease foci that develop into suppurative abscesses. While genetically tractable, the manipulation of S. aureus remains challenging. This unit describes methods developed in our laboratory for gene disruption by allelic replacement and transposition. We also provide a protocol for bacteriophage-mediated transduction of mutants marked with selectable alleles and describe plasmid utilization for complementation studies.

SUBMITTER: Schneewind O

PROVIDER: S-EPMC6249557 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Genetic manipulation of Staphylococcus aureus.

Schneewind Olaf O Missiakas Dominique D

Current protocols in microbiology 20140206

Staphylococcus aureus is a facultative anaerobic Gram-positive coccus and a member of the normal skin flora, as well as that of the nasal passages of humans. However, S. aureus can also gain entry into the host and cause life-threatening infections or persist as disease foci that develop into suppurative abscesses. While genetically tractable, the manipulation of S. aureus remains challenging. This unit describes methods developed in our laboratory for gene disruption by allelic replacement and ...[more]

PMID: 24510849

Similar Datasets

Manipulation of Autophagy in Phagocytes Facilitates Staphylococcus aureus Bloodstream Infection.

Project description:The capacity for intracellular survival within phagocytes is likely a critical factor facilitating the dissemination of Staphylococcus aureus in the host. To date, the majority of work on S. aureus-phagocyte interactions has focused on neutrophils and, to a lesser extent, macrophages, yet we understand little about the role played by dendritic cells (DCs) in the direct killing of this bacterium. Using bone marrow-derived DCs (BMDCs), we demonstrate for the first time that DCs can effectively kill S. aureus but that certain strains of S. aureus have the capacity to evade DC (and macrophage) killing by manipulation of autophagic pathways. Strains with high levels of Agr activity were capable of causing autophagosome accumulation, were not killed by BMDCs, and subsequently escaped from the phagocyte, exerting significant cytotoxic effects. Conversely, strains that exhibited low levels of Agr activity failed to accumulate autophagosomes and were killed by BMDCs. Inhibition of the autophagic pathway by treatment with 3-methyladenine restored the bactericidal effects of BMDCs. Using an in vivo model of systemic infection, we demonstrated that the ability of S. aureus strains to evade phagocytic cell killing and to survive temporarily within phagocytes correlated with persistence in the periphery and that this effect is critically Agr dependent. Taken together, our data suggest that strains of S. aureus exhibiting high levels of Agr activity are capable of blocking autophagic flux, leading to the accumulation of autophagosomes. Within these autophagosomes, the bacteria are protected from phagocytic killing, thus providing an intracellular survival niche within professional phagocytes, which ultimately facilitates dissemination.

| S-EPMC4534639 | biostudies-literature

Genetic diversity of Staphylococcus aureus in Buruli ulcer.

Project description:BackgroundBuruli ulcer (BU) is a necrotizing skin disease caused by Mycobacterium ulcerans. Previous studies have shown that wounds of BU patients are colonized with M. ulcerans and several other microorganisms, including Staphylococcus aureus, which may interfere with wound healing. The present study was therefore aimed at investigating the diversity and topography of S. aureus colonizing BU patients during treatment.MethodologyWe investigated the presence, diversity, and spatio-temporal distribution of S. aureus in 30 confirmed BU patients from Ghana during treatment. S. aureus was isolated from nose and wound swabs, and by replica plating of wound dressings collected bi-weekly from patients. S. aureus isolates were characterized by multiple-locus variable number tandem repeat fingerprinting (MLVF) and spa-typing, and antibiotic susceptibility was tested.Principal findingsNineteen (63%) of the 30 BU patients tested positive for S. aureus at least once during the sampling period, yielding 407 S. aureus isolates. Detailed analysis of 91 isolates grouped these isolates into 13 MLVF clusters and 13 spa-types. Five (26%) S. aureus-positive BU patients carried the same S. aureus genotype in their anterior nares and wounds. S. aureus isolates from the wounds of seven (37%) patients were distributed over two different MLVF clusters. Wounds of three (16%) patients were colonized with isolates belonging to two different genotypes at the same time, and five (26%) patients were colonized with different S. aureus types over time. Five (17%) of the 30 included BU patients tested positive for methicillin-resistant S. aureus (MRSA).Conclusion/significanceThe present study showed that the wounds of many BU patients were contaminated with S. aureus, and that many BU patients from the different communities carried the same S. aureus genotype during treatment. This calls for improved wound care and hygiene.

| S-EPMC4319846 | biostudies-literature

Genetic Regulation of Metal Ion Homeostasis in Staphylococcus aureus.

Project description:The acquisition of metal ions and the proper maturation of holo-metalloproteins are essential processes for all organisms. However, metal ion homeostasis is a double-edged sword. A cytosolic accumulation of metal ions can lead to mismetallation of proteins and cell death. Therefore, maintenance of proper concentrations of intracellular metals is essential for cell fitness and pathogenesis. Staphylococcus aureus, like all bacterial pathogens, uses transcriptional metalloregulatory proteins to aid in the detection and the genetic response to changes in metal ion concentrations. Herein, we review the mechanisms by which S. aureus senses and responds to alterations in the levels of cellular zinc, iron, heme, and copper. The interplay between metal ion sensing and metal-dependent expression of virulence factors is also discussed.

| S-EPMC7494629 | biostudies-literature

Genetic background affects stability of mecA in Staphylococcus aureus.

Project description:The staphylococcal methicillin resistance determinant, mecA, resides on a mobile genetic element, staphylococcus chromosomal cassette mec (SCCmec). The distribution of SCCmec in nature is limited to relatively few clonal complexes of related methicillin-resistant Staphylococcus aureus (MRSA). We have previously reported that some genetic backgrounds are restrictive of mecA and penicillin-binding protein 2a expression, which could account for the restricted clonal distribution of SCCmec in nature. In this study, we investigate the potential role of the host chromosome in the transformability and expression of mecA in 103 naturally occurring methicillin-susceptible S. aureus clinical isolates. The isolates, which had been genotyped previously by multilocus sequence typing, were classified into one of two mutually exclusive categories based on whether the isolates belonged to "major" MRSA lineages or to "other" lineages that are never or occasionally MRSA. We introduced mecA expressed on the low-copy-number plasmid pYK20 into each MSSA strain and assayed the phenotype of resistance to nafcillin by population analysis to assess the relationship between the stability of mecA expression and genetic background. Strains from the major MRSA lineages were more transformable with pYK20 and better able to maintain the plasmid and express resistance in comparison to strains from other lineages. These data support the hypothesis that the presence of mecA within relatively few clonal complexes is partly due to genetic factors that are permissive of mecA and its gene product.

| S-EPMC1153748 | biostudies-literature

The Staphylococcus aureus Host Cell Genetic Adhesion Network

Project description:Adhesion of Staphylococcus aureus to the host’s skin and mucosae enables asymptomatic colonization and the establishment of infections. This process is facilitated by cell wall-anchored (CWA) adhesins that are covalently attached to the bacterial cell wall and exhibit affinity for host ligands. To gain insight into the cellular factors contributing to adhesion, we developed a sensitive and robust assay to enable the large-scale profiling of S. aureus adhesion to host ligands. Using this assay, we profiled a sequence defined transposon mutant library of the community-associated methicillin-resistant S. aureus (CA-MRSA) isolate USA300, to identify mutant strains with attenuated adhesion to human derived keratin, fibronectin and fibrinogen. Overall, we identified 21 adhesion defective mutants and we used this information to construct ‘The S. aureus Genetic Adhesion Network’. Importantly, we identified a core gene set involved in adhesion to all three host ligands and we delineated unique genetic signatures. In addition, we demonstrate a central requirement for autolysin-mediated daughter cell separation in S. aureus CWA protein-mediated adhesion to host ligands, which contributes to virulence in murine skin and soft tissue infection. This comprehensive study will inform strategies to inhibit S. aureus host cell adhesion, potentially enabling the development of anti-adhesive therapeutics to complement traditional antibacterial chemotherapy.

2023-03-04 | PXD018514 | Pride

Manipulation of Autophagy and Apoptosis Facilitates Intracellular Survival of <i>Staphylococcus aureus</i> in Human Neutrophils.

Project description:Polymorphonuclear neutrophils (PMN) are critical for first line innate immune defence against Staphylococcus aureus. Mature circulating PMN maintain a short half-life ending in constitutive apoptotic cell death. This makes them unlikely candidates as a bacterial intracellular niche. However, there is significant evidence to suggest that S. aureus can survive intracellularly within PMN and this contributes to persistence and dissemination during infection. The precise mechanism by which S. aureus parasitizes these cells remains to be established. Herein we propose a novel mechanism by which S. aureus subverts both autophagy and apoptosis in PMN in order to maintain an intracellular survival niche during infection. Intracellular survival of S. aureus within primary human PMN was associated with an accumulation of the autophagic flux markers LC3-II and p62, while inhibition of the autophagy pathway led to a significant reduction in intracellular survival of bacteria. This intracellular survival of S. aureus was coupled with a delay in neutrophil apoptosis as well as increased expression of several anti-apoptotic factors. Importantly, blocking autophagy in infected PMN partially restored levels of apoptosis to that of uninfected PMN, suggesting a connection between the autophagic and apoptotic pathways during intracellular survival. These results provide a novel mechanism for S. aureus intracellular survival and suggest that S. aureus may be subverting crosstalk between the autophagic and apoptosis pathways in order to maintain an intracellular niche within human PMN.

| S-EPMC7686353 | biostudies-literature

Patient sharing and population genetic structure of methicillin-resistant Staphylococcus aureus.

Project description:Rates of hospital-acquired infections, specifically methicillin-resistant Staphylococcus aureus (MRSA), are increasingly being used as indicators for quality of hospital hygiene. There has been much effort on understanding the transmission process at the hospital level; however, interhospital population-based transmission remains poorly defined. We evaluated whether the proportion of shared patients between hospitals was correlated with genetic similarity of MRSA strains from those hospitals. Using data collected from 30 of 32 hospitals in Orange County, California, multivariate linear regression showed that for each twofold increase in the proportion of patients shared between 2 hospitals, there was a 7.7% reduction in genetic heterogeneity between the hospitals' MRSA populations (permutation P value = 0.0356). Pairs of hospitals that both served adults had more similar MRSA populations than pairs including a pediatric hospital. These findings suggest that concerted efforts among hospitals that share large numbers of patients may be synergistic to prevent MRSA transmission.

| S-EPMC3340079 | biostudies-other

Genetic Adaptation of a Mevalonate Pathway Deficient Mutant in Staphylococcus aureus

Project description:In this study we addressed the question how a mevalonate (MVA)-auxotrophic Staphylococcus aureus ΔmvaS mutant can revert to prototrophy.

2020-05-31 | GSE131722 | GEO

Immunogenicity analysis of Staphylococcus aureus clumping factor A genetic variants.

Project description:The staphylococcal adhesin clumping factor A (ClfA) has a variant amino acid sequence, generating the potential for alterations in epitope structure and immunogenicity of this vaccine candidate. We demonstrated for two recombinant ClfA(40-531) (a slightly truncated version of the fibrinogen-binding domain of ClfA containing amino acids 40 to 531) genetic variants that strain-specific epitopes are immunodominant. This work indicates that immune responses elicited by ClfA may, at least in part, be dependent on the strain of origin of the ClfA.

| S-EPMC3754511 | biostudies-literature

Genetic requirements for Staphylococcus aureus nitric oxide resistance and virulence.

Project description:Staphylococcus aureus exhibits many defenses against host innate immunity, including the ability to replicate in the presence of nitric oxide (NO·). S. aureus NO· resistance is a complex trait and hinges on the ability of this pathogen to metabolically adapt to the presence of NO·. Here, we employed deep sequencing of transposon junctions (Tn-Seq) in a library generated in USA300 LAC to define the complete set of genes required for S. aureus NO· resistance. We compared the list of NO·-resistance genes to the set of genes required for LAC to persist within murine skin infections (SSTIs). In total, we identified 168 genes that were essential for full NO· resistance, of which 49 were also required for S. aureus to persist within SSTIs. Many of these NO·-resistance genes were previously demonstrated to be required for growth in the presence of this immune radical. However, newly defined genes, including those encoding SodA, MntABC, RpoZ, proteins involved with Fe-S-cluster repair/homeostasis, UvrABC, thioredoxin-like proteins and the F1F0 ATPase, have not been previously reported to contribute to S. aureus NO· resistance. The most striking finding was that loss of any genes encoding components of the F1F0 ATPase resulted in mutants unable to grow in the presence of NO· or any other condition that inhibits cellular respiration. In addition, these mutants were highly attenuated in murine SSTIs. We show that in S. aureus, the F1F0 ATPase operates in the ATP-hydrolysis mode to extrude protons and contribute to proton-motive force. Loss of efficient proton extrusion in the ?atpG mutant results in an acidified cytosol. While this acidity is tolerated by respiring cells, enzymes required for fermentation cannot operate efficiently at pH ? 7.0 and the ?atpG mutant cannot thrive. Thus, S. aureus NO· resistance requires a mildly alkaline cytosol, a condition that cannot be achieved without an active F1F0 ATPase enzyme complex.

| S-EPMC5884563 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data